Project Summary This research aims for a deeper and more nuanced understanding of the genetics of adaptation than has been possible to date. While many trait-associated variants have now been detected by genome-wide association studies, very few of these SNPs have been directly connected to adaptive phenotypes, and the genetic interactions that govern whether their effects are visible to selection. Such knowledge is crucial to composing realistic and testable models for how widespread standing genetic variation within populations is funneled through the sieve of natural selection. The evolution of melanism in high altitude Drosophila melanogaster populations offers several critical advantages for this endeavor. First, the species offers key functional genetic and population genomic resources, along with a well-annotated genome. Second, prior molecular and evolutionary studies have provided strong background knowledge on the trait, including a compelling set of candidate genes. Third, the study of recent adaptive evolution between populations of the same species maximizes the utility of genetic mapping, population genetics, and functional comparisons of alleles. These features will allow the dissection of this model adaptive trait in unparalleled detail, yielding insights regarding: 1. the functional nature of causative variants, 2. genetic variability of the adaptive response, 3. the prevalence and molecular logic of epistasis among adaptive variants, 4. roles of cryptic variation in adaptive change, 5. the importance of standing genetic variation in trait evolution. Results of this research will advance basic understanding of the adaptive evolutionary process. It will also inform on the importance of genetic background in assessing the phenotypic impact of genetic variants, a key step in understanding the genetic architecture of complex traits including human disease. Investigation of these critical topics will be bolstered by a profoundly integrative research plan that leverages the investigators' complementary backgrounds to fuse novel molecular experiments, genomic analysis, and statistical inference. This research will identify genetic variants underlying melanic adaptation in Ethiopian D. melanogaster, fusing genomic mapping and variation analysis with transgenic tests to pinpoint causative changes (Aim 1). It will also advance beyond that goal to reveal the complex interactions that modulate the phenotypic impact of causative variants (Aim 2), examining tissue- and population-specific gene regulation, and non-additive interactions among melanic variants. These investigations will provide a critical case study that will clarify the complexity of adaptive trait evolution at molecular and genetic levels.